Alteration in intracellular calcium homeostasis reduces motor neuronal viability expressing mutated Cu/Zn superoxide dismutase through a nitric oxide/guanylyl cyclase cGMP cascade

Abstract

Missense mutations in the human Cu/Zn superoxide dismutase gene (SOD-1) cause many cases of autosomal dominant familial amyotrophic lateral sclerosis (FALS). The accumulation of intracellular calcium is one of the primary mechanisms of motor neuronal degeneration associated with mutations in SOD-1. In order to investigate the effect of various calcium modulators and the SOD-1 mutation on neuronal death, we tested motoneuron-neuroblastoma hybrid (VSC 4.1) cells constitutively expressing human SOD-1 gene with mutations (A4V, G93A) or wild-type. These cells were treated with endogenous calcium releaser (ryanodine, thapsigargin, cyclic ADP-ribose) or calcium mobilizer through cell membrane (4-bromo-calcium ionophore A23187). In particular, calcium ionophore reduced survival in the cells expressing mutant SOD-1. Cell death was associated with increased nitric oxide (NO) generation. This toxicity was attenuated when a nitric oxide synthase (NOS) inhibitor was added. Exogenous NOadministration (S-nitrosoglutathione) also induced cell death. The NO-dependent guanylyl cyclase-cGMP cascade inhibitor protected the mutant cells from the toxic effects of calcium ionophore. Our data suggests that motoneuron degeneration with the SOD-1 mutation may be mediated by calcium dysregulation, particularly by the exogenous calcium influx. This process induces oxidative stress generation that results in motor neuronal death through the guanylyl cyclase-cGMP dependent cascade.

The role of phosphodiesterase isoforms 2, 5, and 9 in the regulation of NO-dependent and NO-independent cGMP production in the rat cervical spinal cord

“…20). Expression of sGC in cultured motor neurons has been suggested by a number of groups (Estévez et al, 2002;Kim et al, 2002). However, we have never observed cGMP-IR in motor neurons above background levels.…”

Calcium-influx increases SOD1 aggregates via nitric oxide in cultured motor neurons

“…Moreover, it was found that calcium influx via calcium ionophore stimulated NO production by calmodulin-dependent constitutive NOS (Markram and Segal, 1991). Previously, we showed that calcium ionophore increased NO generation significantly more than endogenous calcium release agents in motor neuronal cells expressing mutant SOD1 (Kim et al, 2002). In the present study, we found that nNOS is involved in calcium influxinduced aggregate formation.…”

Epigallocatechin gallate prevents oxidative-stress-induced death of mutant Cu/Zn-superoxide dismutase (G93A) motoneuron cells by alteration of cell survival and death signals

“…They include oxidative stress, excitotoxicity due to glutamate, calcium-mediated toxicity, genetic defects, autoimmunity, and accumulation of abnormal proteins (Victor and Ropper, 2001). However, increasing evidence indicates that oxidative stress is an important mechanism implicated in neurodegenerative diseases (Al-Chalabi and Leigh, 2000;Kim et al, 2002;Kim et al, 2003;McCord, 1994;Rosen et al, 1993;Son et al, 2003;Sung et al, 2002;Strange et al, 2003;Wang et al, 2002). And, even the precise mechanisms of oxidative stress are not completely characterized, there are growing interests in establishing therapeutic and dietary strategies to protect motor neurons from oxidative-stress-induced damage.…”

“…Although several mechanisms such as enhancement of protein nitrosylation by mutant SOD1, enhanced peroxidase activity, exposure of toxic copper at the active site, and accumulation and aggregation of abnormal proteins have recently been proposed, the precise pathogenic mechanism induced by these mutations has not yet been clearly established (Al-Chalabi and Leigh, 2000;Kim et al, 2002Kim et al, , 2003Kim et al, , 2004McCord, 1994;Rosen et al, 1993;Son et al, 2003;Sung et al, 2002;Strange et al, 2003;Wang et al, 2002).…”